PROJECT SUMMARY/ABSTRACT Chimeric-antigen-receptor T cells (CAR T cells) have achieved extraordinary success in treating B cell malignancies and have demonstrated high potential for treating solid tumors. However, their widespread use is limited by a number of difficulties in manufacturing such cells. Current manufacturing processes are complicated, costly, and can only produce cells for small populations. Additionally, they have large production variations. As a result, different patients receive very different products or treatments. The long-term goal is to develop a novel technology to address the CAR T manufacturing challenge. Toward this goal, the team has developed a proof- of-concept technology termed stress-free intra-tubular cell culture technology (SFIT). SFIT provides cells uniform, highly reproducible, controllable, and cell-friendly microenvironments, resulting in extremely high culture efficiency and consistency. SFIT produces cells for each patient with one small conical tube, significantly reducing production cost and increasing production capability. The purpose of this project is to further develop and validate this technology and the associated methods to make it ready for the final translational challenge. Leveraging sound preliminary studies and a diverse team of experts, the specific aims are to (1) validate SFIT for culturing T cells from a wide range of donors and for culturing various T cell subtypes, (2) further develop and validate methods for transducing T cells with lentivirus and ?-retrovirus in SFIT, and (3) further develop and validate the SFIT-based single-conical-tube-device for producing autologous CAR T cells. The project will build on the team?s strong preliminary work and include mathematical modeling, prototype device design, extensive T cell culturing, exome-Seq and RNA-Seq data analyses, in vivo testing, statistical analyses and comparisons, and partnership with ADS BIOTEC, a company that designs, builds, and sells instruments for processing human cells. With the completion of this project, simple, disposable, affordable devices for the scalable, cost-effective and consistent production of autologous CAR T cells are expected. This technology can also be scaled up for producing allogeneic CAR T cells in large scales if needed. This technology will not only make CAR T cells broadly accessible, but also make the product much more consistent and predictable. Based on 2012-2014 data, approximately 38.5% of the human population will be diagnosed with some type of cancer during their lifetime. Currently, only about 67% of cancer patients will survive five years after diagnosis. CAR T cell therapy is expected to significantly increase their chances of survival. The technology developed in this proposal will make CAR T cells available to many patients.